Engineering a near-infrared dark chromoprotein optimized for photoacoustic imaging
(Conference Presentation)

Yan Li; Quinn Barber; Robert J. Paproski; Jhon R. Enterina; Erik A. Rodriguez; Roger Y. Tsien; Robert E. Campbell; Roger J. Zemp

doi:10.1117/12.2209150

27 April 2016 Engineering a near-infrared dark chromoprotein optimized for photoacoustic imaging (Conference Presentation)

Yan Li, Quinn Barber, Robert J. Paproski, Jhon R. Enterina, Erik A. Rodriguez, Roger Y. Tsien, Robert E. Campbell, Roger J. Zemp

Author Affiliations +

Proceedings Volume 9708, Photons Plus Ultrasound: Imaging and Sensing 2016; 97081Z (2016) https://doi.org/10.1117/12.2209150
Event: SPIE BiOS, 2016, San Francisco, California, United States

Abstract

An optimal genetically-encoded probe for photoacoustic (PA) imaging should exhibit high optical absorption, low fluorescence quantum yield, and an absorption maxima within the near-infrared (NIR) window. One promising candidate is a newly engineered chromoprotein (CP), designated dark small ultra-red fluorescent protein (dsmURFP), which is based on a cyanobacterial phycobiliprotein. To optimize dsmURFP characteristics for PA imaging, we have developed a directed evolution method to iteratively screen libraries of protein variants with three different screening systems. Firstly, we took inspiration from dark-acceptor (also known as dark-quencher)-based Förster resonance energy transfer (FRET) constructs, and used dsmURFP as a dark acceptor from a mCardinal fluorescent donor. The rationale for this design was that the higher the extinction coefficient of the dsmURFP, the more the emission of the donor would be quenched. In addition, more energy transferred to the dark acceptor would lead to more thermoelastic expansion and a stronger PA signal. Three rounds of evolution using this first strategy resulted in dsmURFP1.3 that quenched the emission of mCardinal ~2-fold more efficiently than dsmURFP. Secondly, an absorption-based screening based on visual inspection of plates led to identification of the variant dsmURFP1.4, which exhibited a 2-fold higher absorbance and a 5 nm red shift. Thirdly, we developed a colony-based photoacoustic screening method. To demonstrate the utility of our optimized variants, we used photoacoustic imaging to visualize dsmURFP and its variants in phantom and in vivo experiments using chicken embryo models and murine bacterial bladder infection models.

Conference Presentation

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Yan Li, Quinn Barber, Robert J. Paproski, Jhon R. Enterina, Erik A. Rodriguez, Roger Y. Tsien, Robert E. Campbell, and Roger J. Zemp "Engineering a near-infrared dark chromoprotein optimized for photoacoustic imaging (Conference Presentation)", Proc. SPIE 9708, Photons Plus Ultrasound: Imaging and Sensing 2016, 97081Z (27 April 2016); https://doi.org/10.1117/12.2209150

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KEYWORDS

Photoacoustic imaging

Absorption

Imaging systems

Photoacoustic spectroscopy

Visual process modeling

Fluorescent proteins

Luminescence

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